山羊精液を卵黄緩衝液で稀釈保存すると,多くの場合数日内に白濁凝固して全精子が死滅するが,その際乳酸の蓄積とは無関係に著しくpHの低下することが知られている。また卵黄中には脂質や燐蛋白を多量に含量しており,凝固要因がこれらに作用して酸を生成することは容易に想像される。この実験では手はじめに燐酸や脂肪酸の遊離の有無を検討し,大要次のような結果が得られた。
1. 凝固要因として精漿を使用した場合,凝固時に酸溶性燐と脂質燐に増減が認められ,燐脂質からの部分的な燐酸の遊離が推定されたが,尿道球腺抽出液を凝固要因として使用すると凝固の前後で燐酸量に全く変動がなかつた。このことからこの要因自体には解燐作用はないものと思われ,精漿中に混在する他の要因の作用によるものではないかと思われた。
2. 凝固要因として精漿,尿道球腺抽出液のいずれを使用しても卵黄からの脂肪酸の遊離が認められ,これが凝固時にみられる著しいpHの低下の一因になつているものと思われる。また脂肪酸の遊離にともなつて中性油脂量も増加することが知られた。なお脂肪酸の遊離作用は尿道球腺抽出液の方が精漿に比して約10倍程度強力であつた。

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山羊精液中の卵黄凝固要因の本体を明らかにする目的をもつて,凝固時に遊離される脂肪酸の種類や,凝固要因の卵黄に対する作用位置について検討を加え,大要つぎのような結果が得られた。
1. 卵黄分劃に対する凝固要因の作用を検討することによって卵黄の燐脂質からエステル結合がきれて脂肪酸の遊離されることが知られた。
2. 凝固した卵黄緩衝液について遊離脂肪酸と燐脂質に関するペーパークロマトグラフィーを応用し,その結果遊離される脂肪酸の主なるものはオレイン酸,リノール酸,パルミチン酸などであること,また燐脂質から脂肪酸が遊離されリゾレシチンが生成されることが知られた。
以上の実験結果から,凝固要因は蛇毒性のものと類似のフォスフォリパーゼAに属する酵素と推定された。ただし蛇毒は燐脂質に作用して不飽和酸のみを遊離するのに対して本酵素は飽和酸,不飽和酸の両者を遊離する点で注目すべきものと思われる。

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山羊精液中の卵黄凝固酵素について,その本体とか性質を究明する上に,酵素をできるだけ純粋な型でとり出すことがきわめて重要であり,とくに本酵素作用のごとく新たに知られたものについては,精製の意義は大きいと思われる。本実験では塩析,分別吸着法ならびにアセトン分劃法などによる精製を試みた。得られた主なる結果はつぎのごとくである。
1. 山羊精漿に対して硫安塩析法による分別沈澱を試みたが,硫安濃度30%前後で全蛋白分劃が同時に沈澱し,この方法による酵素の分別は困難であつた。
2. 山羊精漿を塩析,透析後燐酸カルシウムゲルに吸着させ,これを種々の濃度の燐酸緩衝液で分別溶出させることによつて, M/4燐酸緩衝液溶出分劃の酵素純度がかなり上昇することが知られ,精漿に比して純度約10倍の分劃が得られた。
3. 山羊精漿に対してアセトン分劃法を試み,アセトン濃度33%で沈澱する分劃の酵素純度が高く,精漿の約4倍程度に純度を上げることができた。なおアセトンによる分劃を濾紙上に電気泳動させて分別分劃の実態をたしかめ,純度の高い分劃の移動度が低く,ほとんど酵素を含まぬ精のう腺由来と思われる蛋白分劃の移動度の高いことが推定された。

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The biological properties of PMSG are thought to be similar to those of the unfractionated pituitary compound of FSH and ICSH. However, it was proved in the present study that effects of PMSG on immature male reproductive organs is not identical with the pituitary GTH.
The Wistar-Imamichi rats8) weighing 28-40 g at the age of 21-23 days were hypophysectomized through parapharyngeal route. The same assay method reported before6) was used. The results was compared with those obtained by ICSH6) and HCG7) by means of the prostate units (the universal biological unit for comparing the activity of gonadotrophins derived from any source).
Testicular weight increased at the small dose (1.43Pr. U.) of PMSG and plateaued at total dose of about 300 Pr. U. ICSH was slightly effective than PMSG on testicular weight, although ICSH did not give to attain the plateaued weight. Tested by unfractionated sheep pituitary GTH (Vetrophin) however, testicular weight initiated to increase at small dose than ICSH and plateaued on the same level as PMSG at smaller dose (32-64 Pr. U.).
Accordingly, the effect of PMSG to increase the testicular weight of 21 day old rats is weaker than that of the pituitary GTH. In all of these cases, the interstitial cells were stimulated, but the seminiferous tubles were differentiated at only the stage of spermatocyte formation. Accessory organs (epididymides, seminal vesicles and ventral prostate) were extremely stimulated by PMSG as well as HCG. Their plateaued weights were obtained at relatively small doses (about 3 Pr. U. in epididymides and about 20 Pr. U. in seminal vesicles and ventral prostate). By ICSH treatment, the seminal vesicles did not stimulated at higher dose (50 Pr. U.), however, at the more dose of the unfractionated sheep pituitary GTH (about 250 Pr. U.) the weight of seminal vesicles increased to the highest levels in the cases of PMSG and HCG treatment. Similarly, the ventral prostate did not responded conspicuously to ICSH. The weight of ventral prostate of animals received more dose of ICSH (50 Pr. U.) was almost twice of hypophysectomized controls, i. e., the same as animals treated with 1 Pr. U. In this case, higher doses of the unfractionated sheep GTH (about 125-250 Pr. U.) increased the weight of ventral prostate up to the same levels as PMSG and HCG (4-5 times of controls).
It may be said that the distinct difference of PMSG to pituitary GTH is the effect of androgen secretion. The stimulative effect of pituitary GTH to secret the testicular androgen is not conspicuous in immature animals. On the contrary, PMSG having the specific properties of the placental GTH like HCG acceralates powerfully the androgen secretion not only in adult but in immature animals. These differences would be accounted for the intrinsical difference, but not the varied rate of content of FSH and ICSH.

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Ovulation inducing activity of unfractionated sheep pituitary gonadotrophin (Vetrophin) was compared with that of HCG.
One rat ovulating unit3) of Vetrophin was 1/26.7 R. U. by subcutaneous administration and 1/256 R. U. by intravenous injection. The ovulation inducing potency of intravenous injection was about 9.5 times active than that of subcutaneous injection. The ratio of effectiveness of subcutaneous and intravenous injection is almost the same as the results obtained by HCG6).
The potency of Vetrophin is shown by Rat Unit which is determined by the ovarian weight stimulating effects. However, based on the rat ovulating unit, 1 R. U. of Vetrophin coresponds to about 180 to 280 I. U. or 60 to 70 M. U. of HCG by subcutaneous injection and to about 370 I. U. of a preparation of HCG by intravenous injection.
It is said that in terms of HCG tested on the increment of ovarian weight in immature rats, Vetrophin provides the equivalent of 2, 000 I. U. in the 1 R. U. dose8, 11). However, compared by the ovulation inducing activity, 1 R. U. of Vetrophin was equivalent to only about 200 or 400 I. U. of HCG by subcu-taneous or intravenous administration respectively. Vetrophin is used mainly by the intravenous injection for the treatment of the reproductive disorders of domestic animals. On the contrary, HCG is usually not administered intravenously. Considering our results, it should be said that the effect of 1 R. U. of Vetrophin (intravenous injection) to treat the cystic ovaries may be as same as the effect of about 2, 000 I. U. or about 1, 000 M. U. of HCG (subcutaneous injection). The results presented here will furnish some information to clinical use of Vetrophin. Finally, we suggest that in the treatment of cystic ovaries, Vetrophin may be used effectively to treat the cases that antihormone against HCG is produced by previous injections of HCG.

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